Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus responsible for the ongoing global COVID-19 pandemic, exhibits a complex interaction with the host immune system. While viral replication primarily occurs in the upper and lower airways, the long-term effects of SARS-CoV-2 on immune cells, specifically macrophages (Mac) and natural killer (NK) cells, remain a subject of intense research.
Macrophages and NK Cells in SARS-CoV-2 Infection:
Macrophages and NK cells play pivotal roles as frontline innate effector cells against pathogens. However, dysregulated macrophages, as observed in SARS-CoV-2 infection, can contribute to harmful effects on the host, and the role of NK cells in SARS-CoV-2 immunity remains unclear.
The NHP Model and SARS-CoV-2 Variants
Nonhuman primate models, particularly cynomolgus macaques, serve as valuable tools for studying immune cell functioning and responses in tissues. In this study, 25 cynomolgus macaques were infected with wild-type SARS-CoV-2 (WTM), Omicron BA.1, and Omicron BA.2 variants, with noninfected macaques as controls. Viral RNA loads peaked in nasal and tracheal swabs within the first week, and all macaques tested negative for SARS-CoV-2 RNA by day 21. The study extended up to 18 months post-infection, providing a comprehensive understanding of long-term immune responses.
Long-Term Effects on Macrophages
Analysis of bronchoalveolar lavage fluid (BALF) cells at a median of 221 days post-infection revealed a lasting impact on Mac phenotype. The WTM-infected macaques exhibited a higher frequency of CD45+CD64+ Mac and lower lymphocyte frequencies compared to noninfected controls. These changes correlated with elevated levels of inflammatory cytokines, indicating persistent inflammation. Further characterization of BALF Mac revealed distinct clustering and an alternative activation pattern marked by increased expression of CD206, CD4, CD11c, MHC-E, and IL-10.
Reduced IFN-γ production and viability in response to lipopolysaccharide (LPS) stimulation highlighted altered activation in SARS-CoV-2-infected Mac, particularly in the wild-type infection group.
Impaired Natural Killer Cell Responses
The study also explored NK cell responses, revealing a decrease in NK cell frequency during early SARS-CoV-2 infection, along with an exhaustion phenotype. While NK cells from healthy individuals demonstrated the ability to directly kill infected cells in vitro, those from moderate or severe cases exhibited impaired cytotoxic activity.
The production of interferon-γ (IFN-γ) by NK cells was reduced in some macaques, facilitating resistance to NK cell-mediated killing through upregulation of major histocompatibility complex (MHC)-E on the surface of BALF Mac, promoting the persistence of infected cells.
Discussion
This study provides significant insights into the long-term effects of SARS-CoV-2 infection, utilizing both Wuhan and Omicron variants in a cynomolgus macaque model. The findings shed light on the persistence of the virus in lung-resident macrophages (Mac) and its intricate interplay with natural killer (NK) cells, highlighting potential mechanisms that contribute to viral persistence and immune evasion.
Viral Persistence in Macrophages: The detection of viral RNA in bronchoalveolar lavage fluid (BALF) of infected macaques up to 221 days post-infection (p.i.) is a crucial observation. The study demonstrates that SARS-CoV-2 can establish a long-lasting viral reservoir in Mac, even when undetectable in nasopharyngeal and tracheal swabs. This is consistent with persistent viral reservoirs observed in human infections. Notably, the frequency of persisting virus is higher in macaques than in humans, possibly attributed to the high infection doses used in the study.
Potential Mechanisms of Macrophage Infection: The study explores potential mechanisms of SARS-CoV-2 infection in Mac, including direct infection, phagocytosis of infected alveolar epithelial cells, and antibody-dependent enhancement. The presence of viral proteins and RNA in Mac may signify direct infection or phagocytosis. The study’s observations align with in vitro studies showing SARS-CoV-2 entry into human Mac. The role of Mac as a key repository for dissemination and long-term persistence is highlighted, raising questions about other potential cellular reservoirs in the lung.
In Vitro Production of SARS-CoV-2 in Mac: The study demonstrates that SARS-CoV-2 is actively produced in primary cultures of BALF Mac, providing evidence of ongoing viral replication. Morphological changes in the cells, including membrane ruffling and filiform extensions, suggest cell-to-cell dissemination, consistent with reports on tunneling nanotubes. The presence of virus-induced morphological changes and gene expression alterations underscores the active role of Mac in SARS-CoV-2 propagation.
Transcriptomic Changes in Mac: Transcriptomic analysis reveals increased expression of fibronectin (FN1) and S100A8/S100A9 in cultured BALF Mac. These changes may contribute to observed morphological alterations and are reminiscent of chronic TLR4/RAGE signaling reported in long COVID/Postacute sequelae of SARS-CoV-2 infection (PASC). The upregulation of CD1b and MHC-E in BALF Mac suggests noncanonical antigen presentation pathways, potentially manipulated by SARS-CoV-2.
Role of IFN-γ in Viral Persistence: The study emphasizes the role of interferon-gamma (IFN-γ) in inhibiting SARS-CoV-2 replication in BALF Mac. Reduced IFN-γ expression in WTM and OM, both in Mac and NK cells, highlights a potential mechanism contributing to viral persistence. The upregulation of MHC-E on Mac by IFN-γ, while inhibiting viral replication, concurrently renders infected cells more resistant to NK cell-mediated lysis, posing a dilemma in immune responses.
Spike Protein-Mediated Evasion of NK Cells: The study introduces the V3–11 peptide derived from the spike protein, which binds to MHC-E and inhibits NK cell degranulation activity. This suggests a role for the spike protein in evading the immune response. The observation of adaptive NK cells in some macaques, escaping MHC-E-mediated inhibition, underscores the complexity of the host-virus interaction.
Conclusion: In conclusion, this study significantly advances our understanding of the dynamics of SARS-CoV-2 infection in Mac and its impact on NK cell responses. The persistence of the virus, the potential cell-to-cell dissemination, and the intricate immune evasion strategies employed by SARS-CoV-2 highlight the challenges in developing effective therapeutic interventions. Further exploration of the inducibility of adaptive NK cells and their potential modulation through immunotherapies and vaccines is warranted to enhance our strategies against SARS-CoV-2 and future coronaviruses.
reference link: https://www.nature.com/articles/s41590-023-01661-4
